Induced resistance in rainbow trout, Oncorhynchus mykiss (Walbaum), to gill disease associated with the microsporidian gill parasite Loma salmonae

Branchial xenomas were detected by week 5 and disappeared by week 10 after naive juvenile rainbow trout, held at 14.5 °C, were fed or intubated with Loma salmonae ‐infected gill tissue preparations. Upon re‐challenge with L. salmonae , these fish were protected from disease and branchial xenomas did not develop. Branchial xenomas were never detected in naive fish held at 10 °C and exposed to L. salmonae . When these fish were re‐challenged with L. salmonae at 14.5 °C, they were also protected from the disease. Branchial xenomas also developed after naive fish, held at 14.5 °C, were injected intraperitoneally with a semipurified preparation of fresh spores, but generally did not develop after intraperitoneal injection with a preparation of spores subjected to freezing and thawing before use. Fish that had received fresh spores intraperitoneally were completely resistant to disease when re‐challenged via oral delivery of spores, whereas those that had received frozen spores were incompletely, but significantly, protected from disease compared with naive fish. We conclude that infection with L. salmonae induces strong protection towards the disease upon re‐exposure to spores, and that the protection does not depend on the completion of the parasite's life cycle, thus establishing the basis for further research on vaccine development for this disease.     

Characterization of glycans in the developmental stages of Myxobolus cerebralis (Myxozoa), the causative agent of whirling disease

Glycans and sugar-binding molecules (lectins) form an interactive recognition system, which may enable parasitic organisms to adhere to host cells and migrate into target tissues. The aim of the present study was to analyse surface-associated glycans in the developmental stages of Myxobolus cerebralis (Hofer), the causative agent of whirling disease. A panel of biotin-labelled plant lectins was used to detect a broad spectrum of glycan motifs with high specificity. Binding sites were detected histochemically in the tissue sections of infected rainbow trout, Oncorhynchus mykiss (Walbaum), and infected Tubifex tubifex (Müller), and were characterized by light, fluorescence and transmission electron microscopy. With mannose-specific lectins [Lens culinaris agglutinin, Pisum sativum agglutinin, Canavalia ensiformis agglutinin (LCA, PSA, CanA)] mannose-containing glycans were detected in all the developmental stages and host tissues. No binding sites for galactose-specific lectins were present in M. cerebralis spores but reactivity with host tissues occurred. Diversity in glycans was detected by N-acetyl-D-galactosamine-specific lectins in sporoplasm cells of M. cerebralis and triactinomyxon spores. In the group of lectins with monosaccharide-specificity for N-acetyl-D-glucosamine (GlcNAc), the reactivity of Datura stramonium agglutinin (DSA), Lycopersicon esculentum agglutinin (LEA) and Solanum tuberosum agglutinin (STA) was restricted to polar capsules whereas Griffonia simplicifolia agglutinin II (GSA II) also bound to sporoplasm cells of stages in the fish host but not in those present in infected T. tubifex. Moreover, Triticum vulgaris (wheat germ) agglutinin (WGA) and succinylated WGA indicated the presence of N-acetyl-D-glucosamine polymers in polar capsules. No specificity for spores was observed concerning 'bisected'N-glycans and no reactivity in parasitic stages was observed with the fucose-binding lectin Ulex europaeus agglutinin (UEA) I, Sambucus nigra agglutinin (SNA) (specific for alpha2,6-sialylated glycans) and Maackia amurensis agglutinin (MAAI) (specific for alpha2,3-sialylated glycans). Arachis hypogaea (peanut) agglutinin (PNA), Erythrina cristagalli agglutinin (ECA), GSA I, Sophora japonica agglutinin (SJA), Dolichos biflorus agglutinin (DBA) and GSA II detected reactive sites solely confined to the developmental stages of M. cerebralis and were not reactive in the fish host. These parasite-specific glycans may play a role in the adhesion process of the parasite to fish epidermis prior to infection, but may provide protection to the host by activating the complement system, or stimulating an adaptive immune response as putative antigens.     

Hepatic microsporidiosis of juvenile grey mullet, Chelon labrosus (Risso), due to Microgemma hepaticus gen.nov. sp.nov.

Abstract. Microgemma hepaticus gen.nov. sp.nov. is described from the liver of juvenile grey mullet, Chelon labrosus (Risso). Development occurs within xenomas (diameter 500μm) which have microvillar surfaces, encircling bands of mitochondria and a reticulate hypertrophic nucleus. Vegetative developmental stages, meronts, are plasmodial and divided by plasmotomy. These stages are enclosed by host membranes. Sporogonic stages are free in the cytoplasm and divide by multiple exogenous budding. Uninucleate spores (2·4μm × 4·2μm) possess 7–9 coils of the polar filament and a lamellar polaroplast. Xenomas are associated with liver connective tissue, and cause necrosis of adjacent liver cells in certain circumstances. Host response to infection involves leucocyte infiltration and granuloma formation, with spores being destroyed by repeated macrophage phagocytosis and necrosis and resulting in gradual resolution of the lesion. Although juveniles apparently tolerate large parasite burdens there is some evidence of a contribution by the parasite to stress-related mortality. The transmission of the disease and its potentially high pathogenicity to larval fishes is discussed.     

Normal and aberrant tissue distribution of Loma salmonae (Microspora) within rainbow trout, Oncorhynchus mykiss (Walbaum), following experimental infection at water temperatures within and outside of the xenoma-expression temperature boundaries

Temperatures above 20 °C or below 9 °C interrupt the life cycle of the gill intracellular microsporidian parasite Loma salmonae (Microspora) prior to sporogony, inhibiting the production of xenomas. This study intended to characterize this life-cycle failure. Juvenile rainbow trout, Oncorhynchus mykiss (Walbaum), were experimentally infected with L. salmonae spores, and the effect of water temperature on the progress of infection, as determined by polymerase chain reaction, was compared for fish held at water temperatures of 5, 15 and 21 °C. At 15 °C, parasite DNA was first detected in the heart (3 days post-exposure [PE]), and then in the gills and spleen (2 weeks PE). Branchial xenomas developed by week 4 PE. In contrast, at 5 °C, the arrival of the parasite in the heart was delayed until 7 days PE. However, even though parasite DNA was detected in the gills at 7 days PE, xenomas failed to form in the gill, and by week 4 PE, parasite DNA was no longer detected. In fish held at 21 °C, parasite DNA was detected in the heart, gills and spleen by 3 days post-infection, and similar results were observed at 7 days PE. Xenomas also failed to form in these fish and parasite DNA was no longer detected by week 2 PE. Within the range of temperatures tested in this study, spore germination and delivery of their DNA into the host through the intestinal wall was not blocked by temperature. At 5 or 21 °C, migration to the heart and gills occurred, but at aberrant periods of time. The normal life cycle of L. salmonae may depend on the completion of relatively lengthy, but yet unknown, stages of development within the heart, prior to reaching the gill. This development may be adversely affected by temperature, and explain the temperature limits of this parasite.     

Influence of feeding ratio and size on susceptibility to microsporidial gill disease caused by Loma salmonae in rainbow trout, Oncorhynchus mykiss (Walbaum)

Two trials were designed to quantify the effect of feeding ratio and fish size on the cohabitation transmission of Loma salmonae, the causative agent of microsporidial gill disease (MGD) in salmonids, Oncorhynchus spp. To evaluate the effect of feeding ratio on disease onset, groups of 45 rainbow trout, O. mykiss (Walbaum) (RBT), were fed daily at 1% (low), 2% (medium) or 4% (high) of the fish biomass in the tank. There were three tanks at each feeding level: two tanks were exposed to the pathogen and one was a control. For the second objective, 300 RBT were separated into seven tanks so that the weight classifications were small (17-23 g), medium (32-38 g) and large (57-63 g). Each size class was done in duplicate with one control tank containing medium-sized fish. Separately for each trial, on day 0 post-exposure (PE) five highly infectious RBT were added to each tank (not including the control tanks) to begin the cohabitation exposure period. Beginning on day 21 PE and continuing biweekly until days 70 and 77 PE for the feeding and fish size trials, respectively, each fish was evaluated for visible branchial xenomas to determine disease onset time. Using survival analysis, the survival curves for the low, medium and high feedings were not significantly different from each other. However, there were significant differences amongst the small, medium and large weight classes in the size trial. The median numbers of days to the development of branchial xenomas was 31, 38 and 42 for small, medium and large size fish, respectively. On any given day, a medium or large sized fish had a hazard ratio for developing branchial xenomas of 0.66 and 0.63, respectively, compared with a small fish. In addition to host species and host strain differences, fish size is now considered a host risk factor for the development of MGD.     

Modes of transmission of Glugea plecoglossi (Microspora) via the skin and digestive tract in an experimental infection model using rainbow trout, Oncorhynchus mykiss (Walbaum)

Glugea plecoglossi (Microspora) is a significant cause of economic loss in cultured ayu, Plecoglossus altivelis, in Japan, due to the unsightly appearance of infected fish harbouring xenomas in the body cavity. Modes of transmission of G. plecoglossi via the skin and digestive tract were studied in an experimental infection model using rainbow trout, Oncorhynchus mykiss. Combined with Uvitex 2B and in situ hybridization (ISH) assays, the early development of G. plecoglossi was successfully traced. Following a bath exposure of fish Uvitex 2B-labelled G. plecoglossi spores were observed to attach to microscopic injuries (trypan blue-positive sites) of fish skin, after which ISH-positive sporoplasms were found to invade the epidermis as early as 5 min post-infection (PI), migrating rapidly to the subdermis. It was also shown that G. plecoglossi entering via the skin does not spread into the internal organs but develops into subdermal xenomas. After rainbow trout were exposed to G. plecoglossi spores by oral intubation, spores germinated in the intestinal lumen, followed by penetration of sporoplasms into the gut mucosal epithelium 5 min PI. In vitro trials determining stimulation factors (fish mucus, changes in pH, digestive enzymes) for the extrusion of the polar tube were inconclusive. The present study indicates that skin wounds and the gut epithelium can be portals of entry of G. plecoglossi and that natural infection in fish seems to occur perorally rather than via the skin.     

Interaction of water temperature and challenge model on xenoma development rates for Loma salmonae (Microspora) in rainbow trout, Oncorhynchus mykiss (Walbaum)

This study evaluated the regulatory effects of water temperature on the development of branchial xenomas caused by Loma salmonae using a high-dose per os-challenge model compared with a low-dose cohabitation-challenge model. Approximately 275 juvenile rainbow trout (RBT), Oncorhynchus mykiss, were randomly distributed to six tanks with two tanks each maintained at 11, 15 and 19 degrees C. Fish in one tank from each temperature setting were exposed per os to macerated L. salmonae-infected gill material and fish in the other tank from each temperature setting were exposed to L. salmonae using the cohabitation-challenge model. Fish were monitored for the development of branchial xenomas beginning at day 21 post-exposure. Survival analyses were used to evaluate the effect of water temperature and challenge model on the number of days until the first visible branchial xenoma was detected. The survivor curves for the per os-challenge model revealed that there was at least one significant difference, whereas the cohabitation challenge did not reveal any significant differences amongst the temperature settings. The proportional hazards model revealed a significant interaction between the challenge model used and water temperature. This indicated that the effect of water temperature was different depending on challenge model. Additionally, from the mean xenoma intensities, on average, the per os-challenged fish showed higher xenoma intensity compared with the cohabitation-challenged fish. Overall, the impact of water temperature on disease pathogenesis was greater when the RBT were per os challenged compared with using the cohabitation model.     

The life cycle of Henneguya nuesslini Schuberg & Schroder, 1905 (Myxozoa) involves a triactinomyxon-type actinospore

The life cycle of the histozoic myxozoan parasite Henneguya nuesslini was investigated in two salmonid host species. Naive brown trout, Salmo trutta, and brook trout, Salvelinus fontinalis, were experimentally infected in two trials by triactinomyxon type actinospores from naturally infected Tubifex tubifex. In exposed common carp, Cyprinus carpio, no myxospore production was detected. The parasite formed cysts with mature myxospores in the connective tissue of the fish 102 days post-exposure. The morphology of both actinosporean and myxosporean stages was described by light microscopy and a 1417-bp fragment of the 18S rDNA gene was sequenced. Sequence analysis confirmed the absolute congruence of the two developmental stages and assisted in determining species identity. Host range, tissue specificity and myxospore measurements provided sufficiently distinctive features to confirm species validity and were thus crucial for identification. The triactinomyxon spores had 16 secondary germ cells, unique dimensions, a very opaque sporoplasm matrix and three conspicuously protruding, pyriform polar capsules. This is the first record of a Henneguya sp. life cycle with a triactinomyxon-type actinospore, which suggests a close relationship with the Myxobolus group and a polyphyletic origin of the genus Henneguya.     

Spraguea (Microsporida: Spraguidae) infections in the nervous system of the Japanese anglerfish, Lophius litulon (Jordan), with comments on transmission routes and host pathology

Anglerfish from the genus Lophius are a globally important commercial fishery. The microsporidian Spraguea infects the nervous system of these fish resulting in the formation of large, visible parasitic xenomas. Lophius litulon from Japan were investigated to evaluate the intensity and distribution of Spraguea xenomas throughout the nervous system and to assess pathogenicity to the host and possible transmission routes of the parasite. Spraguea infections in L. litulon had a high prevalence; all fish over 403 mm in standard length being infected, with larger fish usually more heavily infected than smaller fish. Seventy percent of all fish examined had some gross visible sign of infection. The initial site of development is the supramedullary cells on the dorsal surface of the medulla oblongata, where all infected fish have parasitic xenomas. As the disease progresses, a number of secondary sites typically become infected such as the spinal, trigeminal and vagus nerves. Fish with infection in the vagus nerve bundles often have simultaneous sites of infection, in particular the spinal nerves and along the ventral nerve towards the urinary bladder. Advanced vagus nerve infections sometimes form xenomas adjacent to kidney tissue. Spraguea DNA was amplified from the contents of the urinary bladders of two fish, suggesting that microsporidian spores may be excreted in the urine. We conclude that supramedullary cells on the hindbrain are the primary site of infection, which is probably initiated at the cutaneous mucous glands where supramedullary cells are known to extend their peripheral axons. The prevalence of Spraguea infections in L. litulon was very high, and infections often extremely heavy; however, no associated pathogenicity was observed, and heavily infected fish were otherwise normal.     

Cell and tissue reactions of turbot Scophthalmus maximus (L.) to Tetramicra brevifilum gen. n., sp. n. (Microspora)

Abstract. Tetramicra brevifilum gen, n. sp, n. is described from the connective tissues throughout the body musculature of turbot Scophthalmus maximus (L.). Uninucleate spores (4.8·2 μm) are characterized by a large posterior vacuole, electron dense inclusion bodies in cytoplasm and nucleus and short filaments with 3–5 coils (50 μm extruded). Sporogony is apansporoblastic and tetrasporoblastic, within a host vacuole. Features of the xenoma (2 mm diameter) include a microvillous surface layer with microtubules and microfilaments and a reticulate nucleus with numerous nucleoli. Adherence between adjacent xenomas results in composite cysts of various sizes and shapes depending on site, being dendritic in the myomeres, arms extending between the fibres. Effects on the host, other than eventual destruction of the host cell, include displacement of muscle fibres in myomeres with loss of attachment to myocommata. Breakdown of the xenoma with release of spores into the connective tissue matrix was associated with a localized necrosis affecting surrounding muscle fibres, with leucocyte infiltration and collagen deposition. Spores phagocytosed by macrophages were destroyed in vacuoles, some with multi-laminate walls. Experimental infections established by intramuscular injection at marked sites remained localized, which suggested that xenomas afford sufficient scope for spore production without the need of secondary invasion. The swimming efficiency of turbot would be impaired in severe infections leading to lowered growth rates and increased mortality in wild populations due to predation and starvation. Feeding hierarchies exist in turbot and the disease could prove significant in farming conditions.     

Whole body net ion fluxes, plasma electrolyte concentrations and haematology during a Loma salmonae infection in juvenile rainbow trout, Oncorhynchus mykiss (Walbaum)

Loma salmonae infections of salmonids culminate in the development of branchial xenomas and subsequent focal hyperplasia of the lamellar or filament epithelium following xenoma rupture and spore release. The effects of this acute branchial disruption upon net ionic flux rates and plasma electrolyte concentrations were determined in juvenile rainbow trout given an experimental oral exposure to L. salmonae. Mean numbers of branchial xenomas peaked at week 5 post-exposure (PE), which coincided with a reduction in the specific growth rate, although there were no significant differences in mass, length or condition of Loma-exposed fish compared with unexposed controls. Following exposure, negative net whole body Na(+) and K(+) fluxes decreased, whereas net Cl(-) fluxes remained unchanged compared with non-exposed control fish. At week 3 PE during the initial branchial xenoma formation stage, there was a significant negative whole body net K(+) flux in Loma-exposed trout compared with other points during the exposure and subsequent infection. Additionally, Loma-exposed fish had marginally elevated plasma Na(+) and Cl(-) concentrations, whilst K(+) levels remained unchanged, compared with control fish. Although there was a progressive decrease in leucocrit, haematocrit remained unchanged over the course of the Loma exposure and subsequent infection. These results suggest that ionic compensation can occur at the gills during the development of xenomas during exposure to L. salmonae and the resultant infection, therefore allowing defence of plasma electrolyte concentrations, unlike the acute ionic disturbances seen with some other parasitic diseases.     

Tissue reaction of the three-spined stickleback Gasterosteus aculeatus L. to infection with Glugea anomala (Moniez, 1887)

Abstract. Studies on an experimental infection of the stickleback (Gasterosteus aculeatus L.) with the microsporidian Glugea anomala conducted until day 126 post infection confirmed that xenoparasitic formations typical of the intracellular development of this microsporidian were responsible for a marked tissue reaction in the host. After a single oral infection, xenoparasitic formations developed in the subepithelial connective tissue of the glandular part of the stomach. The course of xenoma development was asynchronous. Up to the 'cyst' stage (xenomas filled with mature spores), their growth and development caused a pressure atrophy in the surrounding tissue. Later, the response to the presence of the xenoparasitic formation was the development of a granulomatous inflammatory reaction. Locally, the host responded to the infection by phagocytosis of mature spores by locally derived macrophages. In massive infections, changes occurred in the subepithelial connective tissue of the glandular part of the stomach (often the complete disappearance of tubular glands), which may result in a permanent influence on the function of this organ.     

Chicken-derived IgY recognizes developing and mature stages of Loma salmonae (Microsporidia) in Pacific salmon, Oncorhynchus spp.

A polyclonal IgY preparation, purified from yolk of immunized hens, is shown to be useful for the detection of Loma salmonae in Pacific salmon. Purity of the IgY was confirmed by SDS-PAGE and western blots of yolk protein extracts. Reactivity of the IgY preparation to L. salmonae was determined through indirect-fluorescent antibody tests on purified spores and by immunohistochemistry. Developing xenomas in histological sections of heart and gill tissue of infected chinook salmon (Oncorhynchus tshawytscha) were visualized as early as 4 weeks post-exposure (PE). Non-lethal detection of L. salmonae was obtained by indirect-fluorescent microscopy of acetone-fixed gill smears. The IgY preparation was shown to react with another Loma sp., L. branchialis. In contrast, no reactivity was observed with other microsporidian parasites of fish such as Pseudoloma neurophilia and Glugea anomala. Yolk derived IgY may be useful in the detection of other fish pathogens.     

Loma salmonae-associated growth rate suppression in rainbow trout, Oncorhynchus mykiss (Walbaum), occurs during early onset xenoma dissolution as determined by in situ hybridization and immunohistochemistry

Experimental infection of rainbow trout, Oncorhynchus mykiss (Walbaum), juveniles with Loma salmonae at a water temperature of 15 °C yielded detectable parasite DNA within the gills by week 2 post-exposure (PE) and detectable spore-wall antigen within developing xenomas by week 3 PE, as determined by in situ hybridization and monoclonal antibody (Mab) based immunohistochemistry, respectively. The microsporidian was most commonly located within endothelial cells of lamellar basal channels. Whereas the onset of xenoma formation appeared to be relatively synchronous, as expected from previous studies, xenoma dissolution followed an unexpected biphasic pattern with peaks at weeks 4 and 9 PE. The onset of significant growth rate suppression, at week 4 PE in exposed fish, was temporally associated with the appearance of gill lesions which, in turn, were centred about sites of premature xenoma dissolution. The latter was determined by the detection of spore-wall antigen within lesions. Co-habitant control fish began developing xenomas by week 10, indicating the infective potential of those spores released from the principal fish during early xenoma dissolution. Although infection with L. salmonae significantly affects fish growth rates, the time-course of this suppression is limited, and as an unexpected finding, growth rate recovery commences prior to the infection’s resolution.     

Altered tissue distribution of Loma salmonae: effects of natural and acquired resistance

This study compared and contrasted the fate of the microsporidian Loma salmonae , a branchial pathogen of salmonids of the genus Oncorhynchus , upon exposure of (1) naive susceptible rainbow trout (RT) O. mykiss , (2) naive RT passively immunized with sera from RT previously exposed to L. salmonae , (3) previously exposed and resistant RT and (4) two species believed to be innately resistant to the parasite, Atlantic salmon (AS) Salmo salar and brook trout (BT) Salvelinus fontinalis . The fish were infected per os with viable L. salmonae spores. The infection was followed in the fish by detection of parasite DNA by polymerase chain reaction (PCR) at several times post-infection. Spore germination and intestinal invasion by the parasite occurred in all groups of fish. In the susceptible RT, parasite DNA was detected in the heart by day 3 post-exposure (PE), followed by the gill at 2 weeks PE, whereas visible xenoparasitic complexes (xenomas) were detected by week 4 PE. In the passively immunized RT, the parasite's fate was similar to that of controls, however, its arrival in the heart was delayed by 1 week. A delay was also detected in RT which had been previously exposed to L. salmonae and then recovered from disease. In these resistant fish, the parasite was able to reach the heart by week 3 PE, however, it failed to reach the gill and form xenomas. In AS and BT, the parasite reached the heart and gills quickly, where it remained for 2 weeks before being cleared; xenomas never formed. We speculate that failure to complete the life cycle in AS, BT and resistant fish might be because of interference by the immune system in the development of the parasite, resulting in the absence of disease in these fish.     

Host-parasite interaction of the myxosporeans Kudoa paniformis Kabata & Whitaker, 1981 and Kudoa thyrsites (Gilchrist, 1924) in the muscle of Pacific whiting, Merluccius productus (Ayres): an ultrastructural study

Abstract. Kudoa paniformis and Kudoa thyrsites infect the muscle of Pacific whiting, Merluccius productus , and K. paniformis is known to cause a softened muscle texture after death of the fish. Ultrastructural studies were conducted on the host-parasite interaction between Pacific whiting and these myxosporeans at two stages of their infection: (1) unencapsulated plasmodia that contained mature and developing spores, and (2) encapsulated plasmodia that contained deteriorating spores. Unencapsulated plasmodia of both species had a microvillar border that was closely associated with the host muscle. Species-specific characteristics included the presence of multi-vesicular bodies in plasmodia of K. paniformis whereas the cytoplasm of K. thyrsites plasmodia contained an array of vesicles, microfilaments and microtubules. Differences in the predominant organelles of the plasmodia may be related to the production of different proteolytic enzymes by each species. A host response became apparent only after a plasmodium had replaced the width of an infected muscle fibre. At that time, the plasmodia of both species became encapsulated by a wall of fibroblast-like cells that were connected by numerous desmosomes. Occasional phagocytes were located at the edge of these capsules. Ultrastructural evidence suggests that the encapsulated plasmodia were not proteolytically active. Melanin deposition did not occur. However, encapsulated plasmodia with deteriorating spores contained electron-dense breakdown products.     

The transmission potential of Loma salmonae (Microspora) in the rainbow trout, Oncorhynchus mykiss (Walbaum), is dependent upon the method and timing of exposure

The ability of a parasite to transmit from one fish to another is important in the dissemination of disease. Groups of 25 naive rainbow trout (RBT), Oncorhynchus mykiss (Walbaum), were exposed to Loma salmonae by feeding on the viscera (gills, hearts and spleens) from L. salmonae -infected donor RBT (DRBT) or by cohabitation with infected DRBT. Exposure occurred 3, 7, 11 and 15 weeks after the DRBT were infected. All naive RBT were examined 7 weeks post-exposure (PE) to the DRBT. Naive RBT, exposed to DRBT at week 3 PE, by feeding on viscera or by cohabitation, failed to develop visible branchial xenomas. Cohabiting naive RBT with DRBT, at week 7 PE and week 11 PE, resulted in the development of branchial xenomas. Xenomas failed to develop in naive RBT exposed via cohabitation to week 15 PE DRBT. Naive RBT, exposed by feeding on the viscera of DRBT at week 7 PE, week 11 PE and week 15 PE, developed branchial xenomas. The transmission potential of viscera from L. salmonae -infected DRBT at week 15 and week 20 PE was also examined. Naive RBT, fed with viscera free of visible branchial xenomas, from DRBT at week 15 PE and week 20 PE, developed branchial xenomas by week 7 PE. A polymerase chain reaction (PCR) was used to detect L. salmonae DNA from the water and sediments of a tank of L. salmonae -infected RBT at week 7 PE. The method and timing of exposure of naive fish to L. salmonae -infected fish are important in disease transmission and may be useful in predicting and preventing disease outbreaks in aquaculture.     

Loma salmonae-associated xenoma onset and clearance in rainbow trout, Oncorhynchus mykiss (Walbaum): comparisons of per os and cohabitation exposure using survival analysis

Survival analysis techniques were used to compare experimental exposure methods of Loma salmonae (Microspora) in rainbow trout (RBT) by measuring xenoma onset and clearance time. Twenty‐eight naive RBT were exposed per os (fed L. salmonae spores) and 28 RBT were exposed by cohabitation with 28 L. salmonae‐infected RBT. Exposed fish were examined once every week (7 days) post exposure (PE). For xenoma onset, the median survival time, the time to first appearance of branchial xenomas, was 6 weeks PE for both per os and cohabitation exposure. For xenoma clearance, the median survival time, the time to total clearance of branchial xenomas, was 10 weeks for per os exposure and 12 weeks for cohabitation exposure. There was a significant difference between the survival curves of per os fish compared with cohabited fish for both xenoma onset and xenoma clearance. The incidence rate of xenoma development was greater for per os exposure (0.1745 cases per fish‐week) compared with cohabitation exposure (0.1342 cases per fish‐week). Similarly, the rate of xenoma clearance was greater for per os exposure (0.1028 cases per fish‐week) compared with cohabitation exposure (0.0885 cases per fish‐week). Differences between exposure methods were attributed to differences in the frequency and duration of exposure to spores. Survival analysis was useful for examining the onset and clearance of L. salmonae and may have applications for understanding disease dynamics in aquaculture.     

A mortality and associated tissue reactions of chinook salmon, Oncorhynchus tshawytscha (Walbaum), caused by the microsporidan Loma sp.

Abstract. A systemic infection by the microsporidan Loma sp. became epizootic and resulted in 10% mortality of the population of 337 869 juvenile chinook salmon, Oncorhynchus tshawytscha (Walbaum), from Crooked Creek, Alaska, at Fort Richardson Hatchery, Anchorage, Alaska in the spring of 1980. The microsporidan, forming xenoparasitic cysts (xenomas) of the cell-hypertrophy tumour type, was found in the gills, arteries, bulbus arteriosus, pseudobranch, choroid gland and kidney. Individual spores were found in cartilage, musculature, liver, arteries, kidney, choroid gland, bulbus arteriosus, ventricle and atrium. Tissue alterations included inflammation, necrosis, and occlusion of arteries; degeneration and necrosis of cartilage and musculature of the tail and head; pericarditis of the bulbus arteriosus; and hyperplasia of heart tissues. Severe infections could impair swimming efficiency, reduce growth rate, and increase mortality after release due to starvation and predation. This represents the first report of microsporidiosis in hatchery-reared chinook salmon caused by Loma sp.     

Descriptions, development and pathogenicity of myxozoan (Myxozoa: Myxosporea) parasites of juvenile cyprinids (Pisces: Cyprinidae)

Approximately 5000 young of the year (0+) cyprinids comprising roach, chub, dace, minnow, bleak, bream, barbel and gudgeon were examined histologically for the presence of myxozoan infections. Thirteen myxozoans were identified to species, the majority being Myxobolus spp. In addition, two species of Myxidium and of Sphaerospora were recorded. All organs were examined, with the majority of infections being found in the gills, musculature and kidney. However, isolated spores were occasionally found in other tissues. Whilst roach contained the highest number of myxozoan species, it was chub that showed the greatest host response to sporogonic forms. Data are provided on spore morphology, pathogenic responses and tissue and host specificity of the myxozoans recorded.